CN113791146B - Method for detecting organic phosphate pollutants in food and sample treatment method thereof - Google Patents

Abstract

The invention discloses a detection method of organic phosphate pollutants in food and a sample treatment method thereof. The sample processing method comprises the following steps: filling glass fiber filter paper at the bottom of an extraction tank, sequentially filling diatomite, a purification material, a food sample and diatomite, injecting a first extraction solvent for first accelerated solvent extraction to obtain a first extraction liquid, injecting a second extraction solvent for second accelerated solvent extraction to obtain a second extraction liquid, combining the first extraction liquid and the second extraction liquid, and drying to obtain a processed food sample; the purifying material consists of ethylenediamine-N-propylsilane, florisil and alkaline alumina. The obtained processed food sample is directly detected by one or more of the existing detection methods of the organic phosphate pollutants. The invention has simple operation, high automation degree and accurate detection result, and can be widely popularized and applied.

Description

Detection method and sample processing method of organophosphate pollutants in food

technical field

The invention relates to the technical field of detection of organic phosphate pollutants, in particular to a detection method of organic phosphate pollutants in food and a sample processing method thereof.

Background technique

Organophosphate esters (Organophosphate esters, OPEs) as a class of flame retardants and plasticizers are widely used in various industrial products, such as plastics, building materials, textiles, furniture and electronic components. Organic phosphates are directly added to polymers through physical mixing rather than chemical bonding, so they are easily released into environmental media such as air, water, dust, and sludge. In recent years, organophosphates have been detected in breast milk, serum, urine and various foods. Studies have shown that some organophosphates are considered persistent environmental pollutants. Toxicological studies have shown that some organophosphates have adverse reactions such as carcinogenicity, neurotoxicity, nephrotoxicity, reproductive toxicity, hepatotoxicity and endocrine disturbance after high concentration and long-term exposure.

At present, the pretreatment technologies for the detection of organophosphate pollutants in food mainly include solid-phase extraction, dispersive solid-phase extraction, microwave-assisted extraction, matrix solid-phase dispersive extraction, ultrasonic-assisted extraction and QuEChERS methods, etc. The detection methods mainly include gas chromatography- Tandem mass spectrometry (GC-MS/MS) detection method and liquid chromatography-tandem mass spectrometry (LC-MS/MS) detection method, etc. Appropriate sample processing methods are generally adopted for different samples to reduce the interference of other substances in the sample on the test results. At present, there are few reports on the detection methods and sample processing methods of organophosphates in rice noodles for infants and young children.

When analyzing trace amounts of organophosphates in rice flour, wheat flour and corn flour, co-extracted matrix components such as carbohydrates, lipids, and proteins will affect the detection results. Impurity interference is of great significance for the accurate analysis of trace organophosphates in rice flour, wheat flour and corn flour. In addition, due to the large difference in polarity of various organophosphates, when detecting multi-component organophosphates in rice flour, wheat flour and corn flour at the same time, choosing a suitable extraction solvent is crucial to the extraction efficiency of compounds with a wide polarity range. important. Therefore, it is of great significance to establish an accurate, efficient and highly automated method for the quantitative detection of organophosphates in rice flour, wheat flour and corn flour to ensure the safety of food, especially infant food.

Contents of the invention

Aiming at the deficiencies in the prior art, the invention provides a method for detecting organophosphate pollutants in food and a sample processing method thereof, which are easy to operate and have accurate detection results.

A sample processing method for the detection of organophosphate pollutants in food, comprising the steps of:

The bottom of the stainless steel extraction pool is filled with glass fiber filter paper, and then the first diatomite, purification material, food sample and second diatomite are loaded in sequence, and the first extraction solvent is injected to perform the first accelerated solvent extraction to obtain the first extract. Then injecting the second extraction solvent to carry out the second accelerated solvent extraction to obtain the second extract, combining the first extract and the second extract, and drying to obtain the processed food sample;

The purification material consists of ethylenediamine-N-propylsilane (PSA), florisil and basic alumina.

The food includes one or more of rice flour, wheat flour, corn flour and the like.

The organic phosphate pollutants include trimethyl phosphate (TMP), triethyl phosphate (TEP), tripropyl phosphate (TPP), tributyl phosphate (TBP), tris (2-chloroethyl) phosphate ester (TCEP), tris(2-chloroisopropyl) phosphate (TCPP), triphenyl phosphate (TPPA), tris(2-n-butoxyacetate) phosphate (TBEP), tris(1,3 phosphate) - One or more of dichloro-2-propyl) ester (TDCPP), cresyl diphenyl phosphate (CDPP), tricresyl phosphate (TCP), 2-ethylhexyl diphenyl phosphate (EHDPP), etc. kind.

In order to reduce or avoid the interference of other substances in the sample on the test results, the present invention aims at the characteristics of food samples such as rice flour, wheat flour, corn flour, etc., and uses the present invention to design a specific composition of purification material to absorb impurities and purify the matrix. An accelerated solvent extraction step of the extraction solvent, resulting in a processed food sample suitable for use in methods for the detection of organophosphate contaminants.

The sample processing method of the present invention can effectively extract a variety of organophosphate pollutants in a wide polarity range in food, and the obtained processed food samples can effectively overcome the interference of other substances in food on the detection of organophosphate pollutants, At the same time, the simultaneous detection of various organic phosphates in a wide polarity range in food is realized, and the detection results are accurate.

In order to achieve better invention effect, carry out following optimization:

The mass ratio of the first diatomite, purification material, food sample and second diatomite is 2-3:0.05-0.2:1:2-3. Further preferably, the mass ratio of the first diatomite, the purification material, the food sample and the second diatomite is 2:0.1:1:3. The addition of diatomaceous earth can promote the dispersion of the sample, increase the contact area between the sample and the extraction solvent, and make the extraction more complete. In addition, the first layer of diatomaceous earth can help absorb the moisture in the sample, and the second layer of diatomaceous earth is used to fill the extraction cell, which can reduce the amount of extraction solvent used.

According to the characteristics of food such as rice flour, wheat flour and corn flour, PSA is mainly used to remove sugars, fatty acids, organic acids and some polar pigments, etc. Florisil is mainly used to remove grease and wax, etc. Basic alumina is mainly used It is used to remove some polar compounds, such as amino acids and water-soluble vitamins, etc. The combination of the three is beneficial to the adsorption of impurities and the purification of the matrix in food samples such as rice flour, wheat flour and corn flour. In order to ensure the higher recovery rate of the 12 organic phosphate pollutants of the present invention, the mass ratio of PSA, florisil, and basic alumina in the purification material is preferably 0.5-10:1-10:0.1- 10. Further preferably, the mass ratio of ethylenediamine-N-propylsilane, florisil, and basic alumina in the purification material is 2-2.1:2:0.9-1. In order to facilitate the experimental operation, the mass ratio of PSA, florisil, and basic alumina is preferably 2:2:1.

The first extraction solvent is selected from formicated acetonitrile, wherein the volume percentage of formic acid is 0.5%. In view of the wide range of polarity of various organophosphate pollutants, the present invention uses 0.5% by volume formicated acetonitrile with strong polarity to efficiently extract the target substances with strong polarity.

The second extraction solvent is n-hexane. The second extraction solvent is n-hexane with weak polarity, which can realize efficient extraction of weakly polar targets among 12 kinds of organic phosphates with a wide polarity range.

The two-step accelerated solvent extraction using the first extraction solvent and the second extraction solvent is more effective than the one-step extraction with a mixed extraction solvent, and there is a significant difference between the two.

The conditions for the first accelerated solvent extraction include: an extraction temperature of 60°C-100°C (preferably 80°C), an extraction pressure of 1000psi-2000psi (preferably 1500psi), a static extraction time of 5min-10min (preferably 5min), and an extraction cycle of at least 2 times. From the perspectives of resource saving and easy operation, the heating time is preferably 4min-6min (preferably 5min), and the flushing volume is 40%-60% of the volume of the extraction cell (preferably 40% of the volume of the extraction cell).

The conditions for the second accelerated solvent extraction include: an extraction temperature of 60°C-100°C (preferably 80°C), an extraction pressure of 1000psi-2000psi (preferably 1500psi), a static extraction time of 5min-10min (preferably 5min), and an extraction cycle of at least 2 times. From the perspectives of resource saving and easy operation, the heating time is preferably 4min-6min (preferably 5min), and the flushing volume is 40%-60% of the volume of the extraction cell (preferably 40% of the volume of the extraction cell).

If the flushing volume is too small, the flushing may not be complete, and if it is too large, it may cause the volume of the extract to be too large to be collected, increase the time for subsequent drying operations, and the like.

The conditions of the first accelerated solvent extraction may be the same as or different from the conditions of the second accelerated solvent extraction.

The amount of the first extraction solvent and the second extraction solvent is not particularly limited. Generally, it is advisable to keep the sample completely submerged in the extraction solvent during static extraction, and it is advisable to rinse completely during rinsing.

Both the first accelerated solvent extraction and the second accelerated solvent extraction can use accelerated solvent extraction equipment, such as a commercially available accelerated solvent extraction instrument.

A method for detecting organophosphate pollutants in food, using the processed food sample obtained by the sample processing method as a test sample, and directly using one or more methods for detecting organophosphate pollutants. Specifically include: taking test samples, directly using gas chromatography-tandem mass spectrometry (GC-MS/MS) detection method for organophosphate pollutants, liquid chromatography-tandem mass spectrometry (LC-MS/MS) for organophosphate pollutants ) detection method, etc., to detect organic phosphate pollutants in the test sample by one or more methods. The detection method for organophosphate pollutants may be an existing detection method for organophosphate pollutants, or a detection method with simple parameter adjustment based on the existing detection methods for organophosphate pollutants.

The test sample is preferably reconstituted with aqueous methanol solution, filtered, and injected.

The volume ratio of methanol to water in the methanol aqueous solution is preferably 1:1.

The liquid chromatography-tandem mass spectrometry detection method of organophosphate pollutants includes the ultra-high performance liquid chromatography-tandem mass spectrometry detection method of organophosphate pollutants, which has the advantages of high sensitivity, low detection limit and short analysis time . The chromatographic analysis conditions of the ultra-high performance liquid chromatography-tandem mass spectrometry detection method of the organophosphate pollutants preferably include:

Mobile phase A: methanol; mobile phase B: 2mmol/L ammonium formate aqueous solution containing 0.1% formic acid by volume; gradient elution procedure: mobile phase B is maintained at 80% by volume for 1 min, and then reduced to volume in the next 1 min The percentage is 5% and kept for 2 minutes, and finally directly increased to 80% by volume and kept for 2 minutes, the rest is mobile phase A, and the total elution time is 6 minutes.

Organophosphates are often accompanied by blank contamination during the detection process. In order to avoid blank contamination as much as possible, diatomaceous earth and all glassware are preferably heated at 400°C-500°C (most preferably 450°C) for 4h-6h during the experiment, and then covered with aluminum foil. Cover until use.

The raw materials, reagents, instruments, etc. used in the present invention can all adopt commercially available products.

The present invention has the following beneficial effects:

In order to reduce or avoid the interference of other substances in the sample on the test results, the present invention aims at the characteristics of food samples such as rice flour, wheat flour, corn flour, etc., and uses the present invention to design a specific composition of purification material to absorb impurities and purify the matrix. An accelerated solvent extraction step of the extraction solvent, resulting in a processed food sample suitable for use in methods for the detection of organophosphate contaminants.

The processed food sample obtained by adopting the sample processing method of the present invention can effectively overcome the interference of other substances in the food on the detection of organophosphate pollutants, and simultaneously realize the simultaneous detection of various organophosphates in a wide polarity range in the food.

The sample processing method of the present invention has simple steps, high-efficiency extraction process, can effectively extract various organic phosphate pollutants in a wide polarity range in food, and reduces or avoids the extraction of other interfering substances in the sample.

The processed food samples obtained by adopting the sample processing method of the present invention are used as test samples for instrumental detection of organophosphate pollutants, which has a high degree of automation, can detect large batches of samples, and has high recovery rate and good detection effect.

The invention has the advantages of simple and convenient operation, high degree of automation and accurate detection results, and can be widely popularized and applied.

Description of drawings

Fig. 1 is the figure of influence of the number of extraction cycles of the present invention on the rate of recovery of the target;

Fig. 2 is the figure of influence of extraction temperature of the present invention on target object recovery;

Fig. 3 is the composition ratio of the purification material of the present invention and its impact on the recovery rate of the target; wherein, the three sides of the triangle are respectively the mass ratio coordinates of PSA, florisil, and basic alumina;

Fig. 4 is a diagram showing the influence of different extraction solvents on the recovery rate of the target substance, wherein acidified acetonitrile-n-hexane represents that the present invention adopts formicated acetonitrile and n-hexane to accelerate solvent extraction in two steps.

Detailed ways

The present invention will be described in further detail below in conjunction with specific examples, but the protection scope of the present invention is not limited thereto.

In the following examples:

Organic phosphate pollutants include trimethyl phosphate (TMP), triethyl phosphate (TEP), tripropyl phosphate (TPP), tributyl phosphate (TBP), tris(2-chloroethyl) phosphate ( TCEP), tris (2-chloroisopropyl) phosphate (TCPP), triphenyl phosphate (TPPA), tris (2-n-butoxyacetate) phosphate (TBEP), tris (1,3-di Chloro-2-propyl) ester (TDCPP), cresyl diphenyl phosphate (CDPP), tricresyl phosphate (TCP), 2-ethylhexyl diphenyl phosphate (EHDPP).

Blank rice noodle samples, that is, rice noodle samples that do not contain organic phosphate pollutants.

Diatomaceous earth and all glassware are preferably heated at 450 °C for 5 h during the experiment and then covered with aluminum foil until use.

1mg/L organic phosphate mixed standard solution, that is, the above-mentioned 12 kinds of organic phosphate standard substance concentration are all 1mg/L methanol solution.

The analytical conditions of the ultra-high performance liquid chromatography-tandem mass spectrometry detection method for organophosphate pollutants are as follows:

The ultra-high performance liquid chromatography conditions are: ACQUITY UPLC C18 column (2.1mm×100mm, 1.7μm); column temperature: 40°C; flow rate: 0.2mL/min; injection volume: 5.0μL; mobile phase A: methanol; Phase B: 2 mmol/L ammonium formate aqueous solution containing 0.1% (volume percentage) formic acid. Gradient elution program: keep mobile phase B at 80% volume percent for 1 min, drop to 5% volume percentage in the next 1 min and keep it for 2 minutes, and finally rise directly to 80% volume percent and keep it for 2 minutes, the rest is mobile phase A, the total elution time is 6min.

The mass spectrometry conditions are: ion source mode: electrospray ionization source; detection method: multiple reaction monitoring; scanning method: positive ion scanning; spray voltage: 4500V; ion source temperature: 500°C; air curtain gas pressure: 40psi; atomization gas pressure: 50psi; auxiliary air pressure: 50psi.

The multiple reaction monitoring parameters of the analytes are shown in Table 1.

Table 1 Qualitative ion pairs and quantitative ion pairs of 12 organic phosphates

(1) Number of extraction cycles

Add an appropriate amount of 1 mg/L organic phosphate mixed standard solution to the blank rice flour sample to obtain a sample with a single organic phosphate added at 20 μg/kg. Weigh 1.0g sample respectively, change the number of extraction cycles, the number of extraction cycles of the first accelerated solvent extraction and the second accelerated solvent extraction are respectively 1 time, 2 times, 3 times, experiment under the same conditions of the present invention, specifically include :

After the bottom of the stainless steel extraction pool is filled with glass fiber filter paper, 2g of the first diatomite, 0.1g of purification materials composed of PSA, Florisil and basic alumina (in the purification materials, PSA/Florisilicon The mass ratio of soil/basic alumina is 2:2:1), 1.0g of the rice flour sample to be tested and 3g of the second diatomaceous earth. Inject formicated acetonitrile with a volume percentage of 0.5% formic acid as the first extraction solvent. The heating time is 5 minutes, the extraction temperature is 80°C, the extraction pressure is 1500 psi, the static extraction time is 5 minutes, and the extraction cycle is 1, 2 or 3 times. Under the condition that the volume is 40% of the volume of the extraction cell, the sample is subjected to the first accelerated solvent extraction to obtain the first extract. Then inject the second extraction solvent n-hexane, the heating time is 5min, the extraction temperature is 80°C, the extraction pressure is 1500psi, the static extraction time is 5min, the extraction cycle is 1 time, 2 times or 3 times, and the washing volume is 40% of the extraction pool The sample is subjected to a second accelerated solvent extraction under the condition of volume to obtain a second extract. Combine the first extract and the second extract, blow to dryness with nitrogen, vortex reconstitute the residue with 4 mL of 50% by volume methanol aqueous solution, take the supernatant and centrifuge at 10,000 r/min for 5 min, pass through 0.22 μm polytetrafluoroethylene (PTFE) filter membrane, on-machine detection, the detection method adopts the ultra-high performance liquid chromatography-tandem mass spectrometry detection method of organophosphate pollutants.

The experimental results are shown in Figure 1. The recoveries of 12 organic phosphates changed little when the extraction cycles were 2 and 3, and the recoveries of 7 organic phosphates were higher when the extraction cycles were 3. Therefore, it is preferably at least 2 times, more preferably 3 times.

(2) Extraction temperature

Add an appropriate amount of 1 mg/L organic phosphate mixed standard solution to the blank rice flour sample to obtain a sample with a single organic phosphate added at 20 μg/kg. Weigh 1.0g samples respectively, change the extraction temperature to 60°C, 80°C, 100°C, and 120°C respectively, and carry out experiments under the same conditions of the present invention, specifically including:

After the bottom of the stainless steel extraction pool is filled with glass fiber filter paper, 2g of the first diatomite, 0.1g of purification materials composed of PSA, Florisil and basic alumina (in the purification materials, PSA/Florisilicon The mass ratio of soil/basic alumina is 2:2:1), 1.0g of the rice flour sample to be tested and 3g of the second diatomaceous earth. Inject formicated acetonitrile with a volume percentage of 0.5% formic acid as the first extraction solvent, the heating time is 5 minutes, the extraction temperature is 60°C, 80°C, 100°C or 120°C, the extraction pressure is 1500psi, the static extraction time is 5min, and the extraction cycle is 2 The first accelerated solvent extraction was performed on the sample under the condition that the washing volume was 40% of the volume of the extraction cell to obtain the first extract. Then inject the second extraction solvent n-hexane, the heating time is 5min, the extraction temperature is 60°C, 80°C, 100°C or 120°C, the extraction pressure is 1500psi, the static extraction time is 5min, the extraction cycle is 2 times, and the washing volume is 40 Under the condition of % extraction cell volume, the sample is subjected to the second accelerated solvent extraction to obtain the second extract. The first extract and the second extract were combined, blown to dryness with nitrogen, and the residue was vortexed and reconstituted with 4 mL of 50% methanol aqueous solution by volume percentage, and the supernatant was centrifuged at 10000 r/min for 5 min, passed through a 0.22 μm PTFE filter membrane, and placed on The detection method adopts ultra-high performance liquid chromatography-tandem mass spectrometry detection method of organophosphate pollutants.

The experimental results are shown in Figure 2. When the extraction temperature of the two organic phosphates TBEP and TBP was 60°C, the recovery rate reached the highest value; When the temperature was 80°C, the recovery rate reached the highest value; with the increase of temperature, the recovery rate no longer increased or slightly decreased; only TPPA and TCPP reached the maximum recovery at 100 °C, but they were different from those at 80 °C. In contrast, the change in recovery rate was small. Therefore, from the viewpoint of saving energy, it is preferably 60°C-100°C, more preferably 80°C.

(3) Composition and proportion of purification materials

Add an appropriate amount of 1 mg/L organic phosphate mixed standard solution to the blank rice flour sample to obtain a sample with a single organic phosphate added at 20 μg/kg. Design the ternary mixture formula experiment, choose the enhanced simple mass center mixture design, design 10 points and 10 groups of purification materials with different proportions of PSA, Florisil and basic alumina, and the composition of the purification materials for the 10 groups of experiments The proportions are shown in Table 2 and Figure 3. Take 1.0g sample respectively, change the composition of purification material, carry out the experiment that 10 groups of different purification materials are composed under the same conditions of the present invention, each group includes three parallel experiments, calculate the average value as the result of this group, finally calculate the same group of 12 kinds of The average value of the target object results is the response value of this group, and the experimental results are shown in Figure 3. Specifically include:

After inserting glass fiber filter paper at the bottom of the stainless steel extraction pool, add 2 g of the first diatomite and 0.1 g of purification material in turn (see Table 2 and Figure 3 for the mass ratio of PSA/Florisil/basic alumina in the purification material) ), 1.0g of the rice flour sample to be tested and 3g of the second diatomaceous earth. Inject formicated acetonitrile with a volume percentage of 0.5% formic acid as the first extraction solvent. The heating time is 5 minutes, the extraction temperature is 80°C, the extraction pressure is 1500 psi, the static extraction time is 5 minutes, the extraction cycle is 2 times, and the washing volume is 40% of the extraction pool. The sample is subjected to the first accelerated solvent extraction under the condition of the volume to obtain the first extract. Then inject the second extraction solvent n-hexane, the heating time is 5min, the extraction temperature is 80°C, the extraction pressure is 1500psi, the static extraction time is 5min, the extraction cycle is 2 times, and the flushing volume is 40% of the extraction cell volume. A second accelerated solvent extraction is performed to obtain a second extract. The first extract and the second extract were combined, blown to dryness with nitrogen, and the residue was vortexed and reconstituted with 4 mL of 50% methanol aqueous solution by volume percentage, and the supernatant was centrifuged at 10000 r/min for 5 min, passed through a 0.22 μm PTFE filter membrane, and placed on The detection method adopts ultra-high performance liquid chromatography-tandem mass spectrometry detection method of organophosphate pollutants.

The experimental results were analyzed by variance using the fitting curve of Design Expert 10, and the fitting curve was drawn by StatsoftStatistica 12.0. C+0.1305×B×C+7.46×A×A×B×C+3.25×A×B×B×C-1.45×A×B×C×C, where A, B and C are PSA and Flo respectively The proportion fraction of lithium silica and basic alumina, R is the recovery rate/%. Using LINGO15.0 to calculate the extreme value, the calculation results show that when the mass ratio of PSA: florisil: basic alumina is 2.1:2.0:0.9, the average recovery rate of 12 kinds of organic phosphates is the highest, which is 83%. Therefore, for the convenience of subsequent operations, the preferred mass ratio of PSA: florisil: basic alumina is 2:2:1.

Table 2 The mass ratio of the three materials in the purification materials of 10 groups of experiments

group PSA Florisil Basic alumina 1 0 0 10 2 0 5 5 3 0 10 0 4 5 0 5 5 5 5 0 6 10 0 0 7 6.67 1.67 1.67 8 3.3 3.3 3.3 9 1.67 6.67 1.67 10 1.67 1.67 6.67

(4) The order of adding the extraction solvent

Add an appropriate amount of 1 mg/L organic phosphate mixed standard solution to the blank rice flour sample to obtain a sample with a single organic phosphate added at 20 μg/kg.

After the bottom of the stainless steel extraction pool is filled with glass fiber filter paper, 2g of the first diatomite, 0.1g of purification materials composed of PSA, Florisil and basic alumina (in the purification materials, PSA/Florisilicon The mass ratio of soil/basic alumina is 2:2:1), 1.0g of the rice flour sample to be tested and 3g of the second diatomaceous earth. The extraction solvent n-hexane is injected first, and the sample is accelerated under the conditions of heating time of 5 minutes, extraction temperature of 80°C, extraction pressure of 1500 psi, static extraction time of 5 minutes, extraction cycle of 3 times, and flushing volume of 40% of the extraction cell volume. Solvent extraction to obtain a primary extract, which contains more oil. Then inject formicated acetonitrile with 0.5% volume percentage of formic acid as the extraction solvent, the heating time is 5min, the extraction temperature is 80°C, the extraction pressure is 1500psi, the static extraction time is 5min, the extraction cycle is 2 times, and the washing volume is 40% of the extraction pool volume The sample is subjected to accelerated solvent extraction under certain conditions to obtain a secondary extract, which contains more oil. Due to the high oil content in the primary extract and the secondary extract, the matrix effect is too large, which interferes with the test results.

Example 1

Take a rice flour sample, add an appropriate amount of 1 mg/L organophosphate mixed standard solution until the added amount of each organophosphate is 2 μg/kg, and obtain a rice flour sample to be tested.

After the bottom of the stainless steel extraction pool is filled with glass fiber filter paper, 2g of the first diatomite, 0.1g of purification materials composed of PSA, Florisil and basic alumina (in the purification materials, PSA/Florisilicon The mass ratio of soil/basic alumina is 2:2:1), 1.0g of the rice flour sample to be tested and 3g of the second diatomaceous earth. Inject formicated acetonitrile with a volume percentage of 0.5% formic acid as the first extraction solvent. The heating time is 5 minutes, the extraction temperature is 80°C, the extraction pressure is 1500 psi, the static extraction time is 5 minutes, the extraction cycle is 3 times, and the washing volume is 40% of the extraction pool. The sample is subjected to the first accelerated solvent extraction under the condition of the volume to obtain the first extract. Then inject the second extraction solvent n-hexane, the heating time is 5min, the extraction temperature is 80°C, the extraction pressure is 1500psi, the static extraction time is 5min, the extraction cycle is 2 times, and the flushing volume is 40% of the extraction cell volume. A second accelerated solvent extraction is performed to obtain a second extract. The first extract and the second extract were combined, blown to dryness with nitrogen, and the residue was vortexed and reconstituted with 4 mL of 50% methanol aqueous solution by volume percentage, and the supernatant was centrifuged at 10,000 r/min for 5 min, and passed through a 0.22 μm PTFE filter membrane. On-board detection, the detection method adopts the ultra-high performance liquid chromatography-tandem mass spectrometry detection method of organophosphate pollutants.

Test results: Tris(2-n-butoxyacetate) phosphate, tris(2-chloroethyl) phosphate, tributyl phosphate, triethyl phosphate, trimethyl phosphate, triphenyl phosphate, tri( 2-chloroisopropyl) ester, tripropyl phosphate, cresyl diphenyl phosphate, tris(1,3-dichloro-2-propyl) phosphate, tricresyl phosphate, 2-ethylhexyl diphenyl phosphate The rate of recovery of ester is respectively 91%, 90%, 90%, 78%, 75%, 77%, 83%, 84%, 75%, 81%, 73%, 75%; average rate of recovery is 81%; Standard deviation (RSD) ranged from 1% to 12% (n=5).

Example 2

Except for "take a rice flour sample, add an appropriate amount of 1 mg/L organophosphate mixed standard solution until the amount of a single organophosphate added is 20 μg/kg to obtain a rice flour sample to be tested.", the rest of the operations are the same as in Example 1.

The test results are shown in Figure 4: Tris(2-n-butoxyacetate) phosphate, tris(2-chloroethyl) phosphate, tributyl phosphate, triethyl phosphate, trimethyl phosphate, triphenyl phosphate, Tris(2-chloroisopropyl) phosphate, tripropyl phosphate, cresyl diphenyl phosphate, tris(1,3-dichloro-2-propyl) phosphate, tricresyl phosphate, 2-ethyl phosphate The recovery rate of hexyl diphenyl ester is respectively 86%, 87%, 76%, 79%, 74%, 75%, 73%, 75%, 74%, 79%, 71%, 74%; %; RSD 1%-8% (n=5).

Example 3

Except for "take a rice flour sample, add an appropriate amount of 1 mg/L organophosphate mixed standard solution until the added amount of a single organophosphate is 100 μg/kg, to obtain a rice flour sample to be tested.", the rest of the operations are the same as in Example 1.

Test results: Tris(2-n-butoxyacetate) phosphate, tris(2-chloroethyl) phosphate, tributyl phosphate, triethyl phosphate, trimethyl phosphate, triphenyl phosphate, tri( 2-chloroisopropyl) ester, tripropyl phosphate, cresyl diphenyl phosphate, tris(1,3-dichloro-2-propyl) phosphate, tricresyl phosphate, 2-ethylhexyl diphenyl phosphate The recoveries of esters were 90%, 91%, 87%, 79%, 74%, 75%, 78%, 79%, 72%, 82%, 77%, 71%; the average recovery was 80%; RSD 1%-8% (n=5).

Comparative example 1

Take a rice flour sample, add an appropriate amount of 1 mg/L organophosphate mixed standard solution until the added amount of each organophosphate is 20 μg/kg, and obtain a rice flour sample to be tested.

After the bottom of the stainless steel extraction pool is filled with glass fiber filter paper, 2g of the first diatomite, 0.1g of purification materials composed of PSA, Florisil and basic alumina (in the purification materials, PSA/Florisilicon The mass ratio of soil/basic alumina is 2:2:1), 1.0g of the rice flour sample to be tested, and 3g of the second diatomaceous earth. The injected extraction solvent is made of formic acidified acetonitrile with a volume percentage of 0.5% formic acid and n-hexane mixed at a volume ratio of 1:1 (recorded as: acidified acetonitrile: n-hexane 1:1), the heating time is 5min, and the extraction temperature is 80°C , the extraction pressure is 1500psi, the static extraction time is 5min, the extraction cycle is 3 times, and the sample is extracted under the condition that the flushing volume is 40% of the volume of the extraction cell to obtain the extract. The extract was blown to dryness with liquid nitrogen, and the residue was vortexed and reconstituted with 4 mL of 50% methanol aqueous solution by volume percentage, and the supernatant was centrifuged at 10,000 r/min for 5 min, passed through a 0.22 μm PTFE filter membrane, and tested on the machine. The detection method adopts the ultra-high performance liquid chromatography-tandem mass spectrometry detection method of organophosphate pollutants.

The rate of recovery of each organophosphate is shown in Fig. 4, phosphoric acid tris (2-n-butoxyacetate), phosphoric acid tris (2-chloroethyl) ester, tributyl phosphate, triethyl phosphate, trimethyl phosphate, Triphenyl phosphate, tris(2-chloroisopropyl) phosphate, tripropyl phosphate, cresyl diphenyl phosphate, tris(1,3-dichloro-2-propyl) phosphate, tricresyl phosphate, The recoveries of 2-ethylhexyl diphenyl phosphate were 70%, 70%, 69%, 55%, 48%, 67%, 72%, 72%, 63%, 65%, 51%, 60% respectively; The average recovery was 64%; RSD was 1%-10% (n=5). It can be seen that formicated acetonitrile and n-hexane were used as the extraction solvent to accelerate the solvent extraction in one step, and the recoveries of the eight organic phosphates TBP, TEP, TMP, TPPA, CDPP, TDCPP, TCP and EHDPP were low and could not meet the detection requirements.

Comparative example 2

Take a rice flour sample, add an appropriate amount of 1 mg/L organophosphate mixed standard solution until the added amount of each organophosphate is 20 μg/kg, and obtain a rice flour sample to be tested.

After the bottom of the stainless steel extraction pool is filled with glass fiber filter paper, 2g of the first diatomite, 0.1g of purification materials composed of PSA, Florisil and basic alumina (in the purification materials, PSA/Florisilicon The mass ratio of soil/basic alumina is 2:2:1), 1.0g of the rice flour sample to be tested, and 3g of the second diatomaceous earth. Inject the extraction solvent by mixing formic acidized acetonitrile and methanol at a volume ratio of 0.5% formic acid (recorded as: acidified acetonitrile:methanol 1:1). The heating time is 5 minutes, and the extraction temperature is 80°C. The pressure was 1500psi, the static extraction time was 5min, the extraction cycle was 3 times, and the flushing volume was 40% of the volume of the extraction cell to extract the sample to obtain an extract. The extract was blown to dryness with liquid nitrogen, and the residue was vortexed and reconstituted with 4 mL of 50% methanol aqueous solution by volume percentage, and the supernatant was centrifuged at 10,000 r/min for 5 min, passed through a 0.22 μm PTFE filter membrane, and tested on the machine. The detection method adopts the ultra-high performance liquid chromatography-tandem mass spectrometry detection method of organophosphate pollutants.

The rate of recovery of each organophosphate is shown in Fig. 4, phosphoric acid tris (2-n-butoxyacetate), phosphoric acid tris (2-chloroethyl) ester, tributyl phosphate, triethyl phosphate, trimethyl phosphate, Triphenyl phosphate, tris(2-chloroisopropyl) phosphate, tripropyl phosphate, cresyl diphenyl phosphate, tris(1,3-dichloro-2-propyl) phosphate, tricresyl phosphate, The recoveries of 2-ethylhexyl diphenyl phosphate were 55%, 70%, 55%, 71%, 70%, 41%, 56%, 50%, 31%, 46%, 7%, 11% respectively; The average recovery was 47%; RSD was 1%-10% (n=5). It can be seen that formicated acetonitrile and methanol are used as the extraction solvent to accelerate the solvent extraction in one step, and the recoveries of the nine organic phosphates TBEP, TBP, TPPA, TCPP, TPP, CDPP, TDCPP, TCP and EHDPP are low and cannot meet the detection requirements.

Comparative example 3

Take a rice flour sample, add an appropriate amount of 1 mg/L organophosphate mixed standard solution until the added amount of each organophosphate is 20 μg/kg, and obtain a rice flour sample to be tested.

After the bottom of the stainless steel extraction pool is filled with glass fiber filter paper, 2g of the first diatomite, 0.1g of purification materials composed of PSA, Florisil and basic alumina (in the purification materials, PSA/Florisilicon The mass ratio of soil/basic alumina is 2:2:1), 1.0g of the rice flour sample to be tested, and 3g of the second diatomaceous earth. The injection extraction solvent is composed of dichloromethane and n-hexane mixed at a volume ratio of 1:1 (recorded as: dichloromethane:n-hexane 1:1), the heating time is 5min, the extraction temperature is 80°C, and the extraction pressure is 1500psi. The extraction cycle is 3 times, and the sample is extracted under the condition that the washing volume is 40% of the volume of the extraction cell to obtain an extract. The extract was blown to dryness with liquid nitrogen, and the residue was vortexed and reconstituted with 4 mL of 50% methanol aqueous solution by volume percentage, and the supernatant was centrifuged at 10,000 r/min for 5 min, passed through a 0.22 μm PTFE filter membrane, and tested on the machine. The detection method adopts the ultra-high performance liquid chromatography-tandem mass spectrometry detection method of organophosphate pollutants.

The rate of recovery of each organophosphate is shown in Fig. 4, phosphoric acid tris (2-n-butoxyacetate), phosphoric acid tris (2-chloroethyl) ester, tributyl phosphate, triethyl phosphate, trimethyl phosphate, Triphenyl phosphate, tris(2-chloroisopropyl) phosphate, tripropyl phosphate, cresyl diphenyl phosphate, tris(1,3-dichloro-2-propyl) phosphate, tricresyl phosphate, The recoveries of 2-ethylhexyl diphenyl phosphate were 51%, 66%, 58%, 23%, 4%, 39%, 69%, 60%, 45%, 40%, 58%, 63% respectively; The average recovery was 48%; RSD was 2%-11% (n=5). It can be seen that dichloromethane and n-hexane are used as extraction solvents to accelerate solvent extraction in one step, and the recovery rates of 12 kinds of organic phosphates TBEP, TCEP, TBP, TEP, TMP, TPPA, TCPP, TPP, CDPP, TDCPP, TCP and EHDPP are low, reaching Not meet testing requirements.

As can be seen from Fig. 4, compared with the formicated acetonitrile and n-hexane of 0.5% by volume of formic acid in Example 2, which are extracted twice sequentially, Comparative Example 1 uses 0.5% by volume of formicated acetonitrile/n-hexane (1:1 , v/v) The recoveries of the 12 kinds of organic phosphates once extracted as the extraction solvent are generally low, only the recoveries of TCPP and TPP are equivalent, and the recoveries of the remaining 10 kinds of organic phosphates are significantly lower than those in Example 2 (P<0.05 ,Significant difference). In Comparative Example 2, TPPA, TPP, CDPP, TDCPP, TCP, and EHDPP are relatively weak in polarity, and they are extracted with a more polar extraction solvent volume percentage of 0.5% formicated acetonitrile/methanol (1:1, v/v) The recovery rate is low (all less than or equal to 50%), and the average recovery rate of 12 kinds of organic phosphates is lower than 50%; Compared with Example 2, the recovery rate of 12 kinds of organic phosphates in comparative example 2 is generally low, and 12 kinds of organic phosphates Phosphate ester recovery was significantly lower than that in Example 2 (P<0.05, significant difference). In Comparative Example 3, due to the strong polarity of TEP and TMP, the extraction solvent dichloromethane/n-hexane (1:1, v/v) with weaker polarity is used, and the recovery rate of TEP and TMP is below 30%. , all the other 10 kinds of organic phosphate recovery rates are also on the low side; Compared with embodiment 2, the recovery rates of 12 kinds of organic phosphate esters in comparative example 3 are generally on the low side, and 12 kinds of organic phosphate ester recovery rates are obviously lower than embodiment 2 ( P<0.05, the difference was significant). The comparison results show that, under the condition that the purification material (PSA/florisia/basic alumina) is added in the extraction pool proposed by the present invention, the formicated acetonitrile and normal hexane of formic acid volume percentage of 0.5% are injected successively to carry out two extractions , which can realize efficient and accurate detection of organophosphates in a wide polarity range.

Comparative example 4

In addition to "put glass fiber filter paper at the bottom of the stainless steel extraction tank, then add 2g of the first diatomite, 0.1g of purification material (PSA), 1.0g of the rice flour sample to be tested, and 3g of the second diatomite.", the rest Operation is with embodiment 2.

The results are shown in Figure 3, tris (2-n-butoxyacetate) phosphate, tris (2-chloroethyl) phosphate, tributyl phosphate, triethyl phosphate, trimethyl phosphate, triphenyl phosphate , tris(2-chloroisopropyl) phosphate, tripropyl phosphate, cresyl diphenyl phosphate, tris(1,3-dichloro-2-propyl) phosphate, tricresyl phosphate, 2-ethyl phosphate The recoveries of phenylhexyl diphenyl ester are respectively 88%, 81%, 81%, 50%, 24%, 63%, 87%, 87%, 72%, 86%, 67%, 68%; 71%; RSD 5%-15% (n=3). It can be seen that when PSA is used as a purification material combined with two-step accelerated solvent extraction, the recovery rates of TEP and TMP are less than or equal to 50%, and the recovery rates of TPPA, TCP and EHDPP are low, which cannot meet the detection requirements.

Comparative example 5

In addition to "put glass fiber filter paper at the bottom of the stainless steel extraction tank, then add 2g of the first diatomite, 0.1g of the purification material (Florisil), 1.0g of the rice flour sample to be tested, and 3g of the second diatomite." Except, all the other operations are the same as in Example 2.

The results are shown in Figure 3, tris (2-n-butoxyacetate) phosphate, tris (2-chloroethyl) phosphate, tributyl phosphate, triethyl phosphate, trimethyl phosphate, triphenyl phosphate , tris(2-chloroisopropyl) phosphate, tripropyl phosphate, cresyl diphenyl phosphate, tris(1,3-dichloro-2-propyl) phosphate, tricresyl phosphate, 2-ethyl phosphate The recoveries of phenylhexyl diphenyl ester are respectively 62%, 82%, 72%, 74%, 79%, 62%, 61%, 88%, 63%, 64%, 56%, 60%; 69%; RSD 5%-15% (n=3). It can be seen that when florisil was used as a purification material combined with two-step accelerated solvent extraction, the recoveries of the seven organophosphates TBEP, TPPA, TCPP, CDPP, TDCPP, TCP and EHDPP were low and could not meet the detection requirements.

Comparative example 6

In addition to "put glass fiber filter paper at the bottom of the stainless steel extraction tank, then add 2g of the first diatomite, 0.1g of purification material (basic alumina), 1.0g of the rice flour sample to be tested, and 3g of the second diatomite." Except, all the other operations are the same as in Example 2.

The results are shown in Figure 3, tris (2-n-butoxyacetate) phosphate, tris (2-chloroethyl) phosphate, tributyl phosphate, triethyl phosphate, trimethyl phosphate, triphenyl phosphate , tris(2-chloroisopropyl) phosphate, tripropyl phosphate, cresyl diphenyl phosphate, tris(1,3-dichloro-2-propyl) phosphate, tricresyl phosphate, 2-ethyl phosphate The recoveries of phenylhexyl diphenyl ester are respectively 85%, 74%, 79%, 62%, 67%, 63%, 83%, 85%, 62%, 76%, 71%, 69%; 73%; RSD 5%-15% (n=3). It can be seen that when basic alumina is used as a purification material combined with two-step accelerated solvent extraction, the recoveries of the five organic phosphates TEP, TMP, TPPA, CDPP and EHDPP are low and cannot meet the detection requirements.

Comparative example 7

In addition to "put glass fiber filter paper at the bottom of the stainless steel extraction tank and then load 2g of the first diatomaceous earth, 0.1g of purification materials composed of PSA and florisil (wherein the mass ratio of PSA to florisil Be 1:1), 1.0g rice flour sample to be tested, 3g second diatomite." Except, all the other operations are with embodiment 2.

The results are shown in Figure 3, tris (2-n-butoxyacetate) phosphate, tris (2-chloroethyl) phosphate, tributyl phosphate, triethyl phosphate, trimethyl phosphate, triphenyl phosphate , tris(2-chloroisopropyl) phosphate, tripropyl phosphate, cresyl diphenyl phosphate, tris(1,3-dichloro-2-propyl) phosphate, tricresyl phosphate, 2-ethyl phosphate The recoveries of phenylhexyl diphenyl ester are respectively 82%, 84%, 84%, 50%, 55%, 71%, 81%, 83%, 77%, 84%, 71%, 69%; 74%; relative standard deviation (RSD) 5%-15% (n=3). It can be seen that the mass ratio of PSA to florisil is 1:1 as the purification material combined with two-step accelerated solvent extraction, and the recoveries of the three organophosphates TEP, TMP and EHDPP are low and fail to meet the detection requirements.

Comparative example 8

In addition to "put glass fiber filter paper at the bottom of the stainless steel extraction tank and then load 2g of the first diatomite, 0.1g of purification materials composed of PSA and basic alumina (wherein the mass ratio of PSA to basic alumina is 2 : 1), 1.0g rice flour sample to be tested, the second diatomite of 3g." Except, all the other operations are with embodiment 2.

The results are shown in Figure 3, tris (2-n-butoxyacetate) phosphate, tris (2-chloroethyl) phosphate, tributyl phosphate, triethyl phosphate, trimethyl phosphate, triphenyl phosphate , tris(2-chloroisopropyl) phosphate, tripropyl phosphate, cresyl diphenyl phosphate, tris(1,3-dichloro-2-propyl) phosphate, tricresyl phosphate, 2-ethyl phosphate The recoveries of phenylhexyl diphenyl ester are respectively 74%, 71%, 70%, 31%, 27%, 55%, 63%, 68%, 58%, 66%, 57%, 64%; 59%; relative standard deviation (RSD) between 5% and 15% (n=3). It can be seen that the mass ratio of PSA to basic alumina is 2:1 as the purification material combined with two-step accelerated solvent extraction, and the recovery rates of 9 kinds of organic phosphates TEP, TMP, TPPA, TCPP, TPP, CDPP, TDCPP, TCP and EHDPP Low, can not meet the detection requirements.

Comparative example 9

In addition to "put glass fiber filter paper at the bottom of the stainless steel extraction pool, then load 2g of the first diatomite, 0.1g of purification materials composed of florisil and basic alumina (wherein florisil and alkali The mass ratio of permanent alumina is 2:1), 1.0g rice flour sample to be tested, 3g second diatomite." Except, all the other operations are with embodiment 2.

The results are shown in Figure 3, tris (2-n-butoxyacetate) phosphate, tris (2-chloroethyl) phosphate, tributyl phosphate, triethyl phosphate, trimethyl phosphate, triphenyl phosphate , tris(2-chloroisopropyl) phosphate, tripropyl phosphate, cresyl diphenyl phosphate, tris(1,3-dichloro-2-propyl) phosphate, tricresyl phosphate, 2-ethyl phosphate The recoveries of phenylhexyl diphenyl ester are respectively 82%, 78%, 84%, 46%, 36%, 74%, 68%, 79%, 83%, 88%, 107%, 76%; 75%; relative standard deviation (RSD) was 5%-15% (n=3). It can be seen that the mass ratio of florisil to basic alumina is 2:1 as the purification material combined with two-step accelerated solvent extraction, and the recovery rates of the three organophosphates TEP, TMP and TCPP are low and cannot meet the detection requirements .

When the rice flour samples were replaced with wheat flour or corn flour, the test results were equivalent to those of rice flour. It shows that the sample processing method of the present invention is applicable to one or more food samples of rice flour, wheat flour, corn flour, etc. Any combination of parameters within the scope of the present invention can achieve the effect of the present invention. I won't repeat them here.

Claims (9)

1. A sample processing method for detecting organic phosphate pollutants in food, which is characterized by comprising the following steps:

filling glass fiber filter paper into the bottom of a stainless steel extraction tank, sequentially filling first diatomite, a purification material, a food sample and second diatomite, injecting a first extraction solvent for first accelerated solvent extraction to obtain a first extraction liquid, injecting a second extraction solvent for second accelerated solvent extraction to obtain a second extraction liquid, combining the first extraction liquid and the second extraction liquid, and drying to obtain a processed food sample;

the purifying material consists of ethylenediamine-N-propylsilane, florisil and alkaline alumina;

the food comprises one or more of rice flour, wheat flour and corn flour;

the first extraction solvent is selected from methanoized acetonitrile, and the volume percentage of formic acid in the methanoized acetonitrile is 0.5%;

and the second extraction solvent is n-hexane.

2. The method of claim 1, wherein the mass ratio of the first diatomaceous earth, the purification material, the food sample, and the second diatomaceous earth is 2-3:0.05-0.2:1:2-3.

3. The method of claim 2, wherein the mass ratio of the first diatomaceous earth, the purification material, the food sample, and the second diatomaceous earth is 2:0.1:1:3.

4. the method for processing a sample according to claim 1, wherein the mass ratio of ethylenediamine-N-propylsilane, florisil, basic alumina in the purification material is 0.5 to 10:1-10:0.1-10.

5. The method for processing a sample according to claim 4, wherein the mass ratio of ethylenediamine-N-propylsilane, florisil, basic alumina in the purification material is 2 to 2.1:2:0.9-1.

6. The method of claim 1, wherein the first accelerated solvent extraction conditions comprise: extracting at 60-100deg.C under 1000-2000 psi for 5-10 min; the extraction cycle is at least 2 times;

the conditions of the second accelerated solvent extraction include: extracting at 60-100deg.C under 1000-2000 psi for 5-10 min; the extraction cycle is at least 2 times;

the conditions of the first accelerated solvent extraction are the same as or different from the conditions of the second accelerated solvent extraction.

7. A method for detecting organic phosphate contaminants in food, characterized in that a processed food sample obtained by the sample processing method according to any one of claims 1 to 6 is used as a test sample.

8. The method for detecting organic phosphate contaminants in food according to claim 7, wherein the organic phosphate contaminants in the test sample are detected directly by one or more of gas chromatography-tandem mass spectrometry detection of organic phosphate contaminants and liquid chromatography-tandem mass spectrometry detection of organic phosphate contaminants.

9. The method for detecting organic phosphate contaminants in food according to claim 8, wherein the liquid chromatography-tandem mass spectrometry detection method of organic phosphate contaminants includes an ultra-high performance liquid chromatography-tandem mass spectrometry detection method of organic phosphate contaminants, and chromatographic analysis conditions of the ultra-high performance liquid chromatography-tandem mass spectrometry detection method of organic phosphate contaminants include:

mobile phase a: methanol; mobile phase B: 2mmol/L ammonium formate aqueous solution containing 0.1% formic acid by volume; gradient elution procedure: the mobile phase B is kept at 80% by volume for 1min, and then is reduced to 5% by volume for 2min in the next 1min, and finally is directly increased to 80% by volume for 2min, and the balance is the mobile phase A, and the total elution time is 6min.